The aim of this project has been to understand the molecular mechanisms that control the assembly and folding of the MHC class I molecules. The MHC class I molecules serve as the critical cell surface structures that sample intracellular peptide fragments for recognition by antigen- specific, MHC-restricted T lymphocytes. In this project we have sought to understand in an in vitro system how the components of this three chain structure consisting of MHC class I molecule, synthetic peptide, and the light chain subunit, beta2-microglobulin proceed from linear peptide chains, to a properly folded and biologically active MHC molecule. By engineering single chain analogs of the naturally occuring MHC molecules in which each of the components is part of the same polypeptide chain, we have produced model systems for the efficient in vitro folding of class I molecules. Our results indicate that the MHC class I molecule should be considered as a trimer in which each component chain plays a critical role in the pathway to a proper three-dimensional structure. In addition, evidence for an intermediate structure lacking the peptide component, and serologically and functionally distinguishable from the native trimer, has been acquired. Such studies not only permit us to better understand the MHC class I molecule and approaches for engineering variants of it, but also increase our basic understanding of protein folding and its relationship to protein function in general.